Pulsed electromagnetic field processing is a way of affecting biological cells by means of brief pulses of a strong external electromagnetic field. In pulsed electromagnetic field processing two electrodes are placed in a medium comprising biological cells. Then the pulsed electromagnetic field is applied to the medium. The applied electromagnetic field may affect the membranes and/or the intracellular structures of the cells, depending on the field strength and pulse shape (including the time duration and steepness of the pulse). More specifically, a process known as electroporation may occur, which means that the permeability of the cell membrane and/or of the intracellular membrane increases as a result of the electromagnetic field applied to the medium.
When electroporation occurs at the cell membrane, protein channels in the cell membrane may open, causing a change in ion concentration. This change in ion concentration may result in cell stress. When the applied electromagnetic field strength is such that the voltage over the cell membrane is about a critical threshold, for instance a threshold of 1 Volt, and of relative short duration, the cell will recover itself. This effect may be used, for instance, to locally deliver material (such as genes or drugs) into the cells. Applying higher electromagnetic field strengths causing a higher voltage across the cell membrane, and/or applying field pulses of a longer duration, may result in irreversible damage to the cell instead and may eventually even lead to cell death. This effect may be used for processing food, for instance for the cold-pasteurization of fruit juices, or for inducing apoptosis, i.e. programmed cell death in the treatment of diseases such as cancer.
The critical threshold for affecting the cell membrane of the biological cells in the medium may vary, but typically amounts to about 1 Volt. If the typical size of the cells is about 10 μm, the externally applied electric field strength should be in the order of magnitude of 1 kV/cm. This is a very high field strength and in practice it has been difficult to generate electric fields of sufficient strength in the biological cells to be treated.
Furthermore, since electrodes are to be placed inside the medium to be able to generate therein the pulsed electromagnetic field, the biological cells of the medium should be treated first while being in contact with the electrodes, and can be placed in an object, i.e. can be packed into a container, carton, pack(age), etc., only after the cell treatment has been completed. This may reduce the efficiency of the treatment operation considerably.
Another drawback of using electrodes for generating the pulsed electromagnetic field is that the electrodes may erode over time, meaning that eroded electrode material eventually may end up in the medium to be treated. Also electrochemical reactions may occur at or around the electrodes. Such reactions may result in unwanted reaction residues. If the medium includes foodstuff to be consumed by human beings, this may involve an unacceptable health risk.
It is an object of the present invention to provide a method and device for treating biological cells in an object wherein at least one of the above-identified and/or other drawbacks of the prior art have been removed or at least reduced.
It is a more specific object of embodiments of the invention to provide a method and device wherein biological cells in an object may be treated in a fast and efficient manner.
It is a more specific object of embodiments of the invention to provide a method and device wherein biological cells present inside an object may be treated without the need to place one or more electrodes in the object.
According to a first aspect of the invention a device for treating biological cells in an object, the device comprising:
a single winding coil element;
an electrical generator connected to the single winding coil element, the single winding being configured to be positioned essentially around the object;
wherein the electrical generator is configured to discharge into the single winding coil element so that the single winding coil element generates a short duration pulsed electromagnetic field by magnetic induction in the single winding coil element, the electromagnetic field having a field strength that is sufficiently high to affect, preferably increase the permeability of, cell membranes and/or intracellular membranes of the biological cells contained in the object when in operation the object is placed inside the single winding coil element. The device may be electrodeless and therefore the disadvantages associated by the use of electrodes are not present. Furthermore, by placing the object inside the single winding of the coil element the biological cells in the object may be treated without the presence of any electrical element, such as an electrode, in the object. Contact between the device that generated the electromagnetic field and the biological cells can therefore be avoided. Furthermore, since the electromagnetic field is induced by means of a short duration magnetic field, an electric field with the required field strength of at least 1 kV/cm (for affecting cell membranes) or at least 10 kV/cm (for affecting the intracellular membranes) can be realized within a relatively short period of time. Any electroporation brought about in the cells may be used for different purposes, for instance for introducing drugs into the cells. In order to generate the short duration high strength electromagnetic fields needed to bring about the desired effect on the cell membranes and/or the intracellular membranes, a single winding coil element is used. Using a single winding coil element and/or a coaxially built circuit the self-inductance of the coil element and circuit can be kept relatively low, which means that a short rise time of the current in the coil element may be realised. A self-inductance of several tens of nanoHenrys or less can be achieved easily in this embodiment.
In an embodiment the electromagnetic field is applied electrodelessly to the biological cells in the object. Contamination risks due to erosion of the electrodes which in the case of foodstuffs may lead to unacceptable health risks are eliminated.
In a further embodiment the single winding coil element has a generally circular or oval shape in cross-section. This shape allows to generate a pattern of the electromagnetic field that is suitable for affecting, preferably increasing the permeability, of cell membranes and/or intracellular membranes of the biological cells.
In an embodiment the single winding coil element is essentially cylindrical and defines a receiving space sized to allow the object comprising the biological cells to be removably placed. The electromagnetic field is applied when the coil element is placed around the object containing the biological cells or the object is placed inside the receiving space of the single winding coil element. The cylindrical shape of the single winding enables that the entire content in the object is subjected to the high electromagnetic field generated by the single winding coil element.
In an embodiment the single winding coil element defines a receiving space sized to allow the object to pass through the single winding. The object comprising the biological cells can be moved through a stationary single winding of the coil element thereby being subjected to the electromagnetic field. In a further embodiment the coil element can be moved in such a way that the single winding moves along one or more stationary objects comprising biological cells that pass through the receiving space defined by the single winding coil element. In even a further embodiment the coil element comprises multiple single windings, for example arranged in a one-, two-, or three-dimensional array. Again, one or more objects can be passed through the receiving spaces of the respective single windings and/or the single windings can be moved along the one or more objects.
In an embodiment the single winding coil element is configured to allow placement of the object concentrically in the coil element. This may allow the biological cells contained in the object to be subjected to a relatively uniform electromagnetic field of sufficient field strength to affect the cell membranes and/or the intracellular membranes.
In an embodiment the single winding coil is configured to affect the biological cells within the object without contact between the cells and the single winding coil. The absence of physical contact between the single winding coil and the cells to be treated avoids the abovementioned drawbacks of devices employing electrodes for generating and applying the electromagnetic field.
In an embodiment the electrical generator comprises a capacitor arranged so as to generate an electric field strength inside the single winding coil element of at least 10 kV/cm. At least this value of the electric field strength is required to affect the intracellular membranes of the cells.
In an embodiment the winding of the single winding coil element comprises two terminals defining a gap in between. The width of the gap largely determines the pattern of the electromagnetic field and its ability to affect the cell membranes and/or the intracellular membranes of the biological cells in the object that is placed inside the receiving space defined by the single winding coil.
In an embodiment the electrical generator comprises a power supply, a capacitor and a switching element, the switching element being configured for charging the capacitor in a first stage and discharging the capacitor in a second stage so as to provide a short duration high current in the single winding coil element.
In an embodiment the switching element comprises a multiple-gap spark gap switch, preferably a two-gap spark gap switch, configured to break down at a predefined discharge voltage.
In an embodiment the current rise time is 10 ns or less, preferably 6 ns or less, the rate of change of the current (dI/dt) is at least 100 A/ns, preferably at least 150 A/ns, the amplitude of the current pulse is about 500-2000 A, and/or the self-inductance of a circuit comprising the single winding coil element is several tens of nanoHenrys
The switching element may be formed by a multiple-gap spark gap switch, preferably a two-gap spark gap switch, configured to break down at a predefined discharge voltage. The circuit may be built coaxially so that the self inductance is kept to a minimum. These embodiments make it possible to realize a relatively short time duration of the electromagnetic field. More specifically, the rise time of the current in the single winding coil element can be 10 ns or less, preferably 6 ns or less. Moreover, the rate of change of the current (dI/dt) is at least 100 A/ns, preferably at least 150 A/ns, resulting in an amplitude of the current pulse in the single winding coil element of about 500-2000 A. In embodiments of the invention the total duration of the pulse is about several hundreds of nanoseconds, preferably 300 ns.
In an embodiment the generator is at least partly arranged in an enclosure and is pressurized to a predetermined high pressure value, preferably a pressure of at least about 8 bar. The circuit is built coaxially and kept as small as possible to keep the self inductance low of the circuit. In order to reduce the size of the device, more specifically the mutual distance between the elements (for example the multiple spark gap switch), the generator may at least partly be arranged in an enclosure. The enclosure is then pressurized to a predetermined high pressure value, for instance at least about 8 bar.
According to a further aspect an assembly of the device and an object comprising biological cells is provided, the object being arranged in the single winding coil element. The object may be tissue or any solid containing the biological cells. In other embodiments the object is a holder for holding a medium including said biological cells, for instance foodstuffs.
According to another aspect of the invention a method of treating biological cells in an object is provided, the method comprising:                positioning the object inside a single winding of a single winding coil element connected to an electrical generator;        discharging the electrical generator into the single winding coil element so as to generate a short duration pulsed electromagnetic field by magnetic induction in the single winding coil element, wherein the electromagnetic field strength is sufficiently high to affect cell membranes and/or intracellular membranes of the biological cells contained in the object, preferably sufficiently high to increase the permeability of the cell membranes and/or intracellular membranes.        
Preferably the method is suitable for increasing the cell growth rate of the biological cells, and/or increasing the metabolic activity of the biological cells. Advantageously, the biological cells can remain in the medium in which they are contained. There is no need to transfer them to another medium suitable for conventional electroporation.
In an embodiment the method comprises generating an electric field in the object that is sufficiently high, preferably with an electric field strength inside the single winding coil element of at least 1 kV/cm, to affect, preferably increase the permeability of cell membranes and/or intracellular membranes of, the biological cells contained in the object.
In an embodiment the method comprises in a first stage charging a capacitor and in a second stage discharging the capacitor in a second stage so as to provide a short duration high current in the single winding coil element.
In a further embodiment the method comprises increasing the cell growth rate of the biological cells, and/or increasing the metabolic activity of the biological cells.
According to another aspect of the invention a method is provided comprising applying a pulsed electromagnetic field to the biological cells, the electromagnetic field having a field strength sufficiently high to affect, preferably increase the permeability of, cell membranes and/or intracellular membranes of the biological cells contained in the object, wherein the time duration of applying the field to the cells is kept between a minimum treatment time and a maximum treatment time, the treatment time being selected to increase the cell growth rate of the biological cells. The pulsed electromagnetic field may be applied using one or more of the earlier mentioned single winding coils. However, the method also encompasses embodiments wherein the electromagnetic field is applied in a different way, for instance—but not limited to this—by using electrodes, parallel plates, antennas or similar electromagnetic field generating devices.